This proposal represents an integrated approach for opioid design involving the synthesis of building blocks, their incorporation into peptidomimetic opioids, conformational analysis and biological studies. We have developed a Phase I screening protocol which rapidly provides the in vitro and in rive binding affinities for g-,8-, and K-opioid receptors. We have identified three 5-selective drag candidates that will now be synthesized in multigram quantities. This will allow further characterization of their pharmacology and behavioral profiles in order to determine the suitability of 8-preferring agents as drug candidates. Our new Phase II screening protocol includes assays for (i) systemic activity on several mechanistically distinct behavioral models of nociception; (ii) side effect profiles; (iii) effects upon gastrointestinal motility; (iv) tolerance and morphine cross tolerance; (v) effects upon immune function; and (vi) systemic kinetics. A major continuing synthetic program will focus on the preparation of ct-methyl cysteine containing disulfide and lanthionine enkephalins, amine and oxygen bridged enkephalins and scaffold based opioids to search for new target molecules with increased selectivity and potency. These peptidomimetic opioids will be assayed according to the above noted screening protocols. Using molecular biology we intend to determine 5-receptor-ligand interactions. To this end, we are cloning an 86 amino acid peptide corresponding to the transmembrane region IV, extracellular loop II, and transmembrane region V of the 8-opioid receptor which is responsible for ligand binding. This portion of the 8-receptor will be studied by NMR in bilayer and micelle environments. Selective and potent 15N and 13C isotopically labeled ligands will be added and their bound conformations determined. When successful, this will provide the first direct evidence of the "bioactive" conformation of opioid ligands. With the combination of pharmacology, organic chemistry, and molecular biology the design of new highly potent and selective peptidomimetic opioids will be accomplished. [unreadable] [unreadable]